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Abstract A technique for predicting the secular variation (SV) of the geomagnetic field based on the echo state network (ESN) model is proposed. SV is controlled by the geodynamo process in the Earth’s outer core, and modeling its nonlinear behaviors can be challenging. This study employs an ESN to represent the short-term temporal evolution of the geomagnetic field on the Earth’s surface. The hindcast results demonstrate that the ESN enables us to predict SV for a duration of several years with satisfactory accuracy. It is also found that the prediction is robust to the length of the the training data period. This suggests that the recent features of the SV are important for short-term prediction and that the ESN effectively learns these features. Graphical Abstract

Abstract The cellular origin of clear cell ovarian carcinoma (CCOC), a major histological subtype of ovarian carcinoma remains elusive. Here, we explored the candidate cellular origin and identify molecular subtypes using integrated genomic/epigenomic analysis. We performed whole exome-sequencing, microarray, and DNA methylation array in 78 CCOC samples according to the original diagnosis. The findings revealed that ARID1A and/or PIK3CA mutations were mutually exclusive with DNA repair related genes, including TP53, BRCA1, and ATM. Clustering of CCOC and other ovarian carcinomas (n = 270) with normal tissues from the fallopian tube, ovarian surface epithelium, endometrial epithelium, and pelvic peritoneum mesothelium (PPM) in a methylation array showed that major CCOC subtypes (with ARID1A and/or PIK3CA mutations) were associated with the PPM-lile cluster (n = 64). This cluster was sub-divided into three clusters: (1) mismatch repair (MMR) deficient with tumor mutational burden-high (n = 2), (2) alteration of ARID1A (n = 51), and (3) ARID1A wild-type (n = 11). The remaining samples (n = 14) were subdivided into (4) ovarian surface epithelium-like (n = 11) and (5) fallopian tube-like (considered as high-grade serous histotype; n = 3). Among these, subtypes (1–3) and others (4 and 5) were found to be associated with immunoreactive signatures and epithelial-mesenchymal transition, respectively. These results contribute to the stratification of CCOC into biological subtypes.

Background/Aims Intestinal Behçet disease is typically associated with ileocecal punched-out ulcers and significant morbidity and mortality. Intestinal ultrasound is a noninvasive imaging technique for disease monitoring. However, no previous reports have compared intestinal ultrasound with endoscopic ulcer activity or histopathological findings for intestinal Behçet disease. We evaluated the usefulness of intestinal ultrasound for assessing the activity of ileocecal ulcers in intestinal Behçet disease. Methods We retrospectively compared intestinal ultrasound findings with 73 corresponding endoscopic images and 6 resected specimens. The intestinal ultrasound findings were assessed for 7 parameters (bowel wall thickness, vascularity [evaluated using the modified Limberg score with color Doppler], bowel wall stratification, white-plaque sign [strong hyperechogenic lines or spots], mesenteric lymphadenopathy, extramural phlegmons, and fistulas), and endoscopic ulcer activity was classified into active, healing, and scar stages. Histopathological findings were evaluated by consensus among experienced pathologists. Results Bowel wall thickness (P< 0.001), vascularity (P< 0.001), loss of bowel wall stratification (P= 0.015), and white-plague sign (P= 0.013) were significantly exacerbated in the endoscopic active ulcer stage. Receiver operating characteristic curve analysis revealed that a bowel wall thickness of > 5.5 mm (sensitivity 89.7%, specificity 85.3%) was potentially useful for detecting active lesions. When compared with histopathological findings, an increase in bowel wall thickness reflected the ulcer marginal ridge, and the white-plaque sign reflected the ulcer bottom. Conclusions Intestinal ultrasound is useful for monitoring intestinal ulcer activity in intestinal Behçet disease.

Objective: Craniocervical junction arteriovenous fistula (CCJ-AVF) is a rare disease, and preoperative imaging evaluation is important in developing treatment strategies due to the complexity of the vascular architecture, such as the feeding vessels, shunt points, and draining veins. In the present study, we report a case of hemorrhagic perimedullary CCJ-AVF with multiple feeding vessels in which the common single draining vein was dissected by craniotomy and completely treated. Case presentation: A 71-year-old woman presented with subarachnoid hemorrhage (Hunt & Kosnik, grade II). Cerebrovascular CTA showed no intracranial cerebral aneurysm and findings suggestive of shunt disease at the CCJ. Based on cerebral angiography, the patient was diagnosed with perimedullary CCJ AVF, with the right radicular and the left anterior spinal arteries as feeders which drain into a common dilated coronary vein through a shunt located on the ventral side of the spinal cord and then drain into the ascending and descending anterior spinal veins. Ventricular drainage was performed for acute hydrocephalus, and arteriovenous fistula was dissected electively by craniotomy on day 17 after onset. A clip was applied to the single draining vein, which had been evaluated using preoperative imaging diagnosis. Intraoperative digital subtraction angiography (DSA) confirmed the disappearance of shunt blood flow, and the surgery was completed. The follow-up DSA did not show shunt blood flow, and the patient was transferred to a rehabilitation hospital with a modified Rankin scale score of 2. Conclusion: CCJ-AVF cases often have a complex vascular architecture, and it is important to use various modalities to understand the vascular anatomy in detail and develop an appropriate treatment strategy.

Abstract Long interspersed element 1 (LINE-1) open reading frame 1 protein (ORF1p) expression is a common feature of many cancer types, including high-grade serous ovarian carcinoma (HGSOC). Here, we report that ORF1p is not only expressed but also released by ovarian cancer and primary tumor cells. Immuno-multiple reaction monitoring-mass spectrometry assays showed that released ORF1p is confidently detectable in conditioned media, ascites, and patients’ plasma, implicating ORF1p as a potential biomarker. Interestingly, ORF1p expression is detectable in fallopian tube (FT) epithelial precursors of HGSOC but not in benign FT, suggesting that ORF1p expression in an early event in HGSOC development. Finally, treatment of FT cells with DNA methyltransferase inhibitors led to robust expression and release of ORF1p, validating the regulatory role of DNA methylation in LINE-1 repression in non-tumorigenic tissue.

Abstract Background Non-alcoholic fatty liver disease (NAFLD) is usually asymptomatic and lacks a specific biomarker; therefore, many individuals might remain undiagnosed even with advanced liver fibrosis. The aim of this study was to clarify the prevalence and clinical features of subjects with a high risk of advanced liver fibrosis in the general population, using the Fibrosis-4 (FIB-4) index. Methods We retrospectively investigated 6,087 subjects without known liver disease who had participated in an annual health checkup examination. We analyzed the factors associated with high FIB-4 index (≥ 2.67) using a logistic regression analysis. Results Among the 6,087 subjects, 76 (1.2%) had high FIB-4 index. Multivariate analysis identified hypertension (odds ratio [OR]; 9.040; 95% confidence interval [CI], 4.081–20.024; P

Abstract Background Regulatory T cells (Tregs) play an important role in the antitumor immune response in epithelial ovarian cancer (EOC). To understand the immune-inhibitory networks of EOC, we addressed the association between Tregs and immune checkpoint expression on T cells in the tumor microenvironment of EOC. Methods A total of 41 patients with stage IIIC and IV EOC were included in the analysis. We harvested cells from malignant ascites and investigated them using multi-color flow cytometry. We categorized the Tregs into 3 groups: effector-type Tregs, naïve Tregs and non-Tregs, based on the expression patterns of CD45RA and Foxp3 in CD4+ T cells. Furthermore, the relationships between the expression of various immune checkpoint molecules, such as PD-1, on CD8+ T cells and each of the Treg subtypes was also evaluated. Results The median frequency of naïve Tregs, effector-type Tregs and non-Tregs were 0.2% (0–0.8), 2.0% (0–11.4) and 1.5% (0.1–6.3) in CD4+ T cells of malignant ascites from EOC patients, respectively. A high frequency of effector-type Tregs was associated with high-grade serous carcinoma compared with the other histotypes. Patients with higher proportions of effector-type Tregs showed a trend towards increased progression-free survival. We also demonstrated a correlation between a higher proportion of effector-type Tregs and increased PD-1 expression on CD8+ T cells. In addition, C–C chemokine receptor 4 expression was also observed in effector-type Tregs. Conclusion These data suggest that multiple immune-inhibitory networks exist in malignant ascites from EOC patients, suggesting an approach towards combinational immunotherapies for advanced EOC patients.

Abstract Background Administration of poly (ADP-ribose) polymerase (PARP) inhibitors after achieving a response to platinum-containing drugs significantly prolonged relapse-free survival compared to placebo administration. PARP inhibitors have been used in clinical practice. However, patients with platinum-resistant relapsed ovarian cancer still have a poor prognosis and there is an unmet need. The purpose of this study was to examine the clinical significance of metabolic genes and focal adhesion kinase (FAK) activity in advanced ovarian high-grade serous carcinoma (HGSC). Methods The RNA sequencing (RNA-seq) data and clinical data of HGSC patients were obtained from the Genomic Data Commons (GDC) Data Portal and analysed ( https://portal.gdc.cancer.gov/ ). In addition, tumour tissue was sampled by laparotomy or screening laparoscopy prior to treatment initiation from patients diagnosed with stage IIIC ovarian cancer (International Federation of Gynecology and Obstetrics (FIGO) classification, 2014) at the Saitama Medical University International Medical Center, and among the patients diagnosed with HGSC, 16 cases of available cryopreserved specimens were included in this study. The present study was reviewed and approved by the Institutional Review Board of Saitama Medical University International Medical Center (Saitama, Japan). Among the 6307 variable genes detected in both The Cancer Genome Atlas-Ovarian (TCGA-OV) data and clinical specimen data, 35 genes related to metabolism and FAK activity were applied. RNA-seq data were analysed using the Subio Platform (Subio Inc, Japan). JMP 15 (SAS, USA) was used for statistical analysis and various types of machine learning. The Kaplan-Meier method was used for survival analysis, and the Wilcoxon test was used to analyse significant differences. P < 0.05 was considered significant. Results In the TCGA-OV data, patients with stage IIIC with a residual tumour diameter of 1-10 mm were selected for K means clustering and classified into groups with significant prognostic correlations (p = 0.0444). These groups were significantly associated with platinum sensitivity/resistance in clinical cases (χ2 test, p = 0.0408) and showed significant relationships with progression-free survival (p = 0.0307). Conclusion In the TCGA-OV data, 2 groups classified by clustering focusing on metabolism-related genes and FAK activity were shown to be associated with platinum resistance and a poor prognosis.

Abstract Background Real‐world outcomes of nivolumab treatment for gastric cancer and associated prognostic factors remain unclear; the present study aimed to evaluate both items. Methods A total of 278 consecutive patients treated with nivolumab for gastric cancer during 2017‐2019 were enrolled in this multi‐institutional retrospective cohort study. The impact of laboratory findings, immune‐related adverse events (irAEs), and clinicopathological factors on long‐term survival was evaluated using the Cox proportional hazards model. Results The response rate was 11.7% in patients with measurable lesions. The overall and progression‐free survival estimates were 6.77 and 2.53 months, respectively. The incidence of irAEs was 30.6% (6.8% for grade ≥3). There were no treatment‐related deaths. Multivariate analysis revealed that C‐reactive protein level of ≤0.5 mg/dL (hazard ratio = 0.476, P 3.5 g/dL (hazard ratio = 0.688, P = .045), performance status 0 (hazard ratio = 0.711, P = .028), lymphocyte count >1000/μL (hazard ratio = 0.686, P = .027), and differentiated histological type (hazard ratio = 0.740, P = .046) were independently associated with improved survival. The median survival of patients with four or more good prognostic factors was 18.3 months. Conclusion Nivolumab showed safety and survival benefits in patients with previously treated unresectable or recurrent gastric cancer. Low C‐reactive protein level, irAE occurrence, high albumin level, high lymphocyte count, and differentiated histological type may affect outcomes. The presence of four or more good prognostic factors may help identify likely long‐term survivors.

Abstract Aims Recent advances of direct‐acting antiviral drugs for hepatitis C virus (HCV) have dramatically improved the sustained virologic response (SVR) rate, but hepatocellular carcinoma (HCC) development rarely occurs even in patients who achieve an SVR. Wisteria floribunda agglutinin‐positive mac‐2‐binding protein (WFA+‐M2BP) was recently developed as a noninvasive biomarker of liver fibrosis. However, the association between the WFA+‐M2BP level and HCC development after the achievement of an SVR is unclear. Methods and Results We examined the association between WFA+‐M2BP and HCC development in 522 HCV patients who achieved an SVR (Interferon [IFN]‐based therapy, n = 228; IFN‐free therapy, n = 294). Multivariate analysis revealed that a high WFA+‐M2BP level at SVR week 24 after treatment (SVR24) (hazard ratio [HR] = 1.215, P = 0.020), low platelet counts (HR = 0.876, P = 0.037), and old age (HR = 1.073, P = 0.012) were independent risk factors for HCC development regardless of the treatment regimen. Receiver operator characteristics curve analysis revealed that a WFA+‐M2BP level at SVR24 of ≥1.62 cut‐off index (COI) was the cut‐off value for the prediction of HCC development (adjusted HR = 12.565, 95% CI 3.501–45.092, P

Abstract Accumulation of tau protein is a key pathology of age-related neurodegenerative diseases such as Alzheimer's disease and progressive supranuclear palsy. Those diseases are collectively termed tauopathies. Tau pathology is associated with axonal degeneration because tau binds to microtubules (MTs), a component of axon and regulates their stability. The acetylation state of MTs contributes to stability and histone deacetylase 6 (HDAC6) is a major regulator of MT acetylation status, suggesting that pharmacological HDAC6 inhibition could improve axonal function and may slow the progression of tauopathy. Here we characterize N-[(1R,2R)-2-{3-[5-(difluoromethyl)-1,3,4-oxadiazol-2-yl]-5-oxo-5H,6H,7H-pyrrolo[3,4-b]pyridin-6-yl}cyclohexyl]-2,2,3,3,3-pentafluoropropanamide (T-518), a novel, potent, highly selective HDAC6 inhibitor with clinically favorable pharmacodynamics. T-518 shows potent inhibitory activity against HDAC6 and superior selectivity over other HDACs compared with the known HDAC6 inhibitors in the enzyme and cellular assays. T-518 showed brain penetration in an oral dose and blocked HDAC6-dependent tubulin deacetylation at Lys40 in mouse hippocampus. A 2-week treatment restored impaired axonal transport and novel object recognition in the P301S tau Tg mouse, tauopathy model, while a 3-month treatment also decreased RIPA-insoluble tau accumulation. Pharmaceutical inhibition of HDAC6 is a potential therapeutic strategy for tauopathy, and T-518 is a particularly promising drug candidate.

Abstract Gaucher disease, the most prevalent metabolic storage disorder, is caused by mutations in the glucocerebrosidase gene GBA1, which lead to the accumulation of glucosylceramide (GlcCer) in affected cells. Gaucher disease type 1 (GD1), although defined as a nonneuronopathic subtype, is accompanied by an increased risk of Parkinson's disease. To gain insights into the association of progressive accumulation of GlcCer and the Parkinson's disease phenotypes, we generated dopaminergic (DA) neurons from induced pluripotent stem cells (iPSCs) derived from a GD1 patient and a healthy donor control, and measured GlcCer accumulation by liquid chromatography‐mass spectrometry. We tested two DA neuron differentiation methods: a well‐established method that mimics a step‐wise developmental process from iPSCs to neural progenitor cells, and to DA neurons; and a synthetic mRNA‐based method that overexpresses a transcription factor in iPSCs. GD1‐specific accumulation of GlcCer was detected after 60 days of differentiation by the former method, whereas it was detected after only 10 days by the latter method. With this synthetic mRNA‐based rapid differentiation method, we found that the metabolic defect in GD1 patient cells can be rescued by the overexpression of wild‐type GBA1 or the treatment with an inhibitor for GlcCer synthesis. Furthermore, we detected the increased phosphorylation of α‐synuclein, a biomarker for Parkinson's disease, in DA neurons derived from a GD1 patient, which was significantly decreased by the overexpression of wild‐type GBA1. These results suggest that synthetic mRNA‐based method accelerates the analyses of the pathological mechanisms of Parkinson's disease in GD1 patients and possibly facilitates drug discovery processes.

Methods: Two hundred and forty seven of 480 patients with naïve papilla undergoing therapeutic ERCP between April 2013 and March 2018 were enrolled for the study. The following patient characteristics were investigated: age, sex, body mass index, previous diseases (heart disease, renal failure, cerebrovascular disorders, coexisting malignancy and pulmonary disease), history of PEP, common bile duct diameter, diverticula and volume of fluid infused 24 hours after the procedure. All ERCP cases had naïve papilla and had undergone treatment. Results: The incidence of PEP was 8.5%. Significant differences were observed in the volume of fluid infused between patients without and with a history of heart disease (1,380 vs. 1,755 mL). The mean volume of the infused fluid was significantly lower in the PEP than non-PEP group (1,483 vs. 1,688 mL, P = 0.02). Moreover, PEP incidence differed according to a fluid infusion cutoff of 1,000 mL (7 vs. 11 cases of PEP in those with ≦1,000 mL and >1,000 mL fluid volume, respectively, P < 0.001). Conclusion: Restricted fluid volume was a newly identified risk factor for PEP, particularly in patients with heart and renal diseases as comorbidities.

A 54-year-old woman complained about a headache. Six days later, she developed subcortical hemorrhage (4 × 4 × 5 cm) due to reversible cerebral vasoconstriction syndrome (RCVS). She underwent a craniotomy procedure and maintained oral administration of verapamil. She suffered from higher brain dysfunction, was transferred to convalescent rehabilitation hospital at mRS3 stage, and recovered to mRS0 stage in three months. However, because there was cognitive decline such as inability to calculate and because there was a ventricular enlargement on the image, we performed an LP shunt procedure under the judgment of secondary hydrocephalus. Her cognitive function has improved, and the postoperative course is going smoothly.

Low-frequency noise may create specific perceptions, which might cause various health effects. The present study aimed to identify exposure–response relationships between low-frequency noise and perceptions by re-analysing an experimental study. We investigated the predominant perceptions of ‘feeling bothered’ and ‘feeling of pressure and/or vibration’ using multivariate logistic regression analysis. A significant interaction between 1/3 octave-band sound pressure level and frequency was indicated for ‘feeling bothered’ but not ‘feeling of pressure and/or vibration’, suggesting that the ‘feeling of pressure and/or vibration’ does not originate in cochlear. A mathematical model indicating resonance at approximately 50 Hz fitted the results well. A frequency weighting derived from the mathematical model could be applied to broad-band low-frequency noise to evaluate the perception and health effects. However, further investigations on the weighting for the perception are necessary because the results were obtained only from the existing study.

Background: Patients taking low-dose aspirin have a higher incidence of gastroduodenal ulcers and higher risk of upper gastrointestinal bleeding than patients who do not. Thienopyridine antiplatelet agents may similarly cause bleeding gastroduodenal erosions and ulcers. The incidence of gastrointestinal bleeding is reported to be higher when these antithrombotic drugs are used in combination. Until now, most studies have focused on bleeding, and no study has compared the degree of gastric mucosal injury between patients receiving dual antiplatelet therapy (DAPT) and those receiving single antiplatelet therapy (SAPT) in real-world clinical practice. Aim: Our objective was to compare the degree of gastric mucosal injury in patients taking low-dose aspirin in combination with clopidogrel (one of the thienopyridine antiplatelet agents) with that of patients who were taking aspirin or clopidogrel as a single agent. Methods: Patients who were taking aspirin and/or clopidogrel and who underwent scheduled esophagogastroduodenoscopy between April 2015 and March 2020 were enrolled in this study. Endoscopic images were reviewed retrospectively, and the degree of gastric mucosal injury was assessed with the modified Lanza score (m-Lanza score). The m-Lanza score was compared between DAPT patients taking aspirin and clopidogrel and SAPT patients taking either aspirin alone or clopidogrel alone. Results: The m-Lanza scores of the DAPT group, the aspirin group, and the clopidogrel group were 1.67 ± 1.81 (mean ± standard deviation), 0.95 ± 1.61, and 0.72 ± 1.29, respectively. The m-Lanza score of the DAPT group tended to be higher than that of the aspirin group (p = 0.06) and was significantly higher than that of the clopidogrel group (p = 0.03). Conclusion: The degree of gastric mucosal injury in DAPT patients was significantly higher than that in patients using clopidogrel alone and tended to be higher than that in patients using aspirin alone in real-world clinical practice.

Oncolytic vaccinia virus (OVV) has demonstrated appropriate safety profiles for clinical development. Although designed to kill cancer cells efficiently, OVV sensitivity varies in individual cancers, and predictive biomarkers of therapeutic responses have not been identified. Here we found that OVV was much more efficient in KFTX paclitaxel-resistant ovarian cancer cells compared to that in KFlow paclitaxel-sensitive cells. Microarray analysis identified long non-coding RNA urothelial carcinoma-associated 1 (UCA1) upregulation, which contributed to both enhanced paclitaxel resistance and OVV spread. In addition, UCA1 expression correlated with efficient OVV spread in other ovarian cell lines and primary cancer cell cultures. When host pathways underlying OVV spread were analyzed, differences were detected in the activation of the Rho GTPase Cdc42, suggesting that filopodia formation enhances OVV cell-to-cell spread and tumor migration. Moreover, we established a clinically relevant mouse model of peritoneal metastasis using KFTX or KFlow cells. Paclitaxel exerted anti-tumor effects on KFlow, but not KFTX, tumors. In mice bearing KFTX cells after paclitaxel failure, OVV treatment induced the regression of residual tumors and improved survival. Our findings demonstrated that UCA1 promotes OVV cell-to-cell spread in ovarian cancer, resulting in enhanced therapeutic outcome. Keywords: oncolytic vaccinia virus, biomarker, long non-coding RNA, UCA1, chemotherapy

Pancreatic intraepithelial neoplasia (PanIN), the most common premalignant lesion of the pancreas, is a histologically well-defined precursor to invasive pancreatic ductal adenocarcinoma (PDAC). However, the molecular mechanisms underlying the progression of PanINs have not been fully elucidated. Previously, we demonstrated that the expression of collapsin response mediator protein 4 (CRMP4) in PDAC was associated with poor prognosis. The expression of CRMP4 was also augmented in a pancreatitis mouse model. However, the role of CRMP4 in the progression of PanIN lesions remains uncertain. In the present study, we examined the relationship between CRMP4 expression and progression of PanIN lesions using genetically engineered mouse models. PanIN lesions were induced by peritoneal injection of the cholecystokinin analog caerulein in LSL-KRASG12D; Pdx1-Cre (KC-Crmp4 wild-type, WT) mice and LSL-KRASG12D; Pdx1-Cre; Crmp4−/− (KC-Crmp4 knockout, KO) mice. We analyzed pancreatic tissue sections from these mice and evaluated PanIN grade by hematoxylin and eosin staining. CRMP4 expression was examined and the cellular components assessed by immunohistochemistry using antibodies against CRMP4, CD3, and α-smooth muscle actin (SMA). The incidence of high-grade PanIN in KC-Crmp4 WT mice was higher than that in KC-Crmp4 KO animals. CRMP4 was expressed not only in epithelial cells but also in αSMA-positive cells in stromal areas of PanIN lesions. The CRMP4 expression in stromal areas correlated with PanIN grade in WT mice. These results suggested that the expression of CRMP4 in stromal cells may underlie the incidence or progression of PanIN.

We reported previously that EGAM1 homeoproteins (EGAM1 and EGAM1N), transcribed from the Crxos gene as splicing variants, are expressed in preimplantation mouse embryos and mouse embryonic stem (ES) cells. Exogenous expression of these proteins affects the maintenance of an undifferentiated state and the progression of differentiation in mouse ES cells. Human tetrapeptide-repeat homeobox 1 (TPRX1), a member of the eutherian totipotent cell homeobox (ETCHbox) genes, is an ortholog of Crxos. However, the roles of TPRX1 in the differentiation of human pluripotent cells are still unknown. Because the TPRX1 transcripts were undetectable in an undifferentiated state and during the progression of differentiation in wild-type human embryonal carcinoma NT2/D1 cells, it would be advantageous to clarify the relationship between the exogenous expression of TPRX1 and the induction of genes encoding lineage-specific transcription factors in pluripotent cells. The expression of GATA6 and FOXA2, crucial transcription factors for the formation of the primitive endoderm, was upregulated, whereas that of CDX2, a crucial transcription factor for the formation of the trophectoderm, was downregulated by enforced expression of TPRX1. Overall, we suggest that TPRX1 is capable of modulating the expression of lineage-specific transcription factors in pluripotent cells derived from humans.

In this paper, we propose an image-sensor-based visible light communication (IS-VLC) system that provides visual information in digital signage used for advertisements. One important issue of IS-VLC systems is the deterioration of visual information while transmitting the data information. Human eyes are sensitive to luminance but not to color difference; therefore, we propose the modulation of color difference. We evaluate the performance of the proposed system in terms of both the quality of visual information and communication quality of data information, and clarify the effectiveness of the proposed system.

We aimed to analyze the serum level of a novel fibrosis marker, Mac-2-binding protein glycosylation isomer (M2BPGi), and its predictive value for hepatocellular carcinoma (HCC) development in chronic hepatitis B (CHB) under nucleot(s)ide analogue (NA) therapy. Serum M2BPGi levels were quantified in 147 CHB patients at baseline, 48 weeks after starting NA therapy, and at the patients’ last visit. The serum M2BPGi level serially decreased at each time point. During the median follow-up time of 6.6 years, 14 of 147 patients developed HCC. Multivariate Cox proportional hazard analysis demonstrated that high serum M2BPGi at 48 weeks was an independent risk factor for HCC development. A cutoff value of M2BPGi at 48 weeks > 1.5 showed an adjusted hazard ratio = 34.9 (95% confidence interval, 4.3−284.9). The 3- and 5-year cumulative incidence of HCC in patients with low M2BPGi were 0.9% and 4.2%, respectively, whereas those in patients with high M2BPGi were 10.1% and 25.6%, respectively (p < 0.001). In conclusion, Serum M2BPGi level at 48 weeks is a useful predictor for HCC development in patients with CHB who receive NA therapy.

Viral vectors that can infect neurons transsynaptically and can strongly express foreign genes are useful for investigating the organization of neural circuits. We previously developed a propagation-competent rabies virus (RV) vector based on a highly attenuated HEP-Flury strain (rHEP5.0-CVSG), which selectively infects neurons and propagates between synaptically connected neurons in a retrograde direction. Its relatively low level of transgene expression, however, makes immunostaining necessary to visualize the morphological features of infected neurons. To increase the transgene expression level of this RV vector, in this study we focused on two viral proteins: the large protein (L) and matrix protein (M). We first attempted to enhance the expression of L, which is a viral RNA polymerase, by deleting the extra transcription unit and shortening the intergenic region between the G and L genes. This viral vector (rHEP5.0-GctL) showed increased transgene expression level with efficient transsynaptic transport. We next constructed an RV vector with a rearranged gene order (rHEP5.0-GML) with the aim to suppress the expression of M, which plays a regulatory role in virus RNA synthesis. Although this vector showed high transgene expression level, the efficiency of transsynaptic transport was low. To further evaluate the usability of rHEP5.0-GctL as a transsynaptic tracer, we inserted a fluorescent timer as a transgene, which changes the color of its fluorescence from blue to red over time. This viral vector enabled us the differentiation of primary infected neurons from secondary infected neurons in terms of the fluorescence wavelength. We expect this propagation-competent RV vector to be useful for elucidating the complex organization of the central nervous system.

Objective: Ovarian cancer has the highest mortality among gynecological cancers. High-grade serous carcinoma (HGSC) is the most common histotype of ovarian cancer regardless of ethnicity, whereas clear cell carcinoma (CCC) is more common in East Asians than Caucasians. The elucidation of predominant signaling pathways in these cancers is the first step towards understanding their molecular mechanisms and developing their clinical management. Methods: RNA sequencing was performed for 27 clinical ovarian specimens from Japanese women. Principal component analysis (PCA) was conducted on the sequence data mapped on RefSeq with normalized read counts, and functional annotation analysis was performed on genes with substantial weights in PCA. Knockdown experiments were conducted on the selected genes on the basis of PCA. Results: Functional annotation analysis of PCA-defined genes showed predominant pathways, such as cell growth regulators and blood coagulators in CCC and transcription regulators in HGSC. Knockdown experiments showed that the inhibition of the calcium-dependent protein copine 8 (CPNE8) and the transcription factor basic helix-loop-helix family member e 41 (BHLHE41) repressed the proliferation of CCC- and HGSC-derived cells, respectively. Conclusions: This study identified CPNE8 and BHLHE41 as characteristic genes for CCC and HGSC, respectively. The systemic identification of differentially expressed genes in CCC and HGSC will provide useful information to understand transcriptomic differences in these ovarian cancers and to further develop potential diagnostic and therapeutic options for advanced disease.

The glycoprotein-gene (G gene) -deleted rabies virus (RV) vector is a powerful tool to examine the function and structure of neural circuits. We previously reported that the deletion of the G gene enhances the transgene expression level of the RV vector. However, the mechanism of this enhancement remains to be clarified. We presume that there are two possible factors for this enhancement. The first factor is the glycoprotein of RV, which shows cytotoxicity; thus, may cause a dysfunction in the translation process of infected cells. The second possible factor is the enhanced expression of the L gene, which encodes viral RNA polymerase. In the RV, it is known that the gene expression level is altered depending on the position of the gene. Since G-gene deletion displaces the L gene in the genome, the expression of the L gene and viral transcription may be enhanced. In this study, we compared the transgene expression level and viral transcription of three recombinant RV vectors. The effect of glycoprotein was examined by comparing the viral gene expression of G-gene-intact RV and G-gene-replaced RV. Despite the fact that the L-gene transcription level of these two RV vectors was similar, the G-gene-replaced RV vector showed higher viral transcription and transgene expression level than the G-gene-intact RV vector. To examine the effect of the position of the L gene, we compared the viral gene expression of the G-gene-deleted RV and G-gene-replaced RV. The G-gene-deleted RV vector showed higher L-gene transcription, viral transcription, and transgene expression level than the G-gene-replaced RV vector. These results indicate that G-gene deletion enhances the transgene expression level through at least two factors, the absence of glycoprotein and enhancement of L-gene expression. These findings enable investigators to design a useful viral vector that shows a controlled desirable transgene expression level in applications.

The aim of this study is to determine if ultraviolet light (UVC) irradiation in combination with fluorescence-guided surgery (FGS) can eradicate metastatic human pancreatic cancer in orthotopic nude-mouse models. Two weeks after orthotopic implantation of human MiaPaCa-2 pancreatic cancer cells, expressing green fluorescent protein (GFP), in nude mice, bright-light surgery (BLS) was performed on all tumor-bearing mice (n = 24). After BLS, mice were randomized into 3 treatment groups; BLS-only (n = 8) or FGS (n = 8) or FGS-UVC (n = 8). The residual tumors were resected using a hand-held portable imaging system under fluorescence navigation in mice treated with FGS and FGS-UVC. The surgical resection bed was irradiated with 2700 J/m2 UVC (254 nm) in the mice treated with FGS-UVC. The average residual tumor area after FGS (n = 16) was significantly smaller than after BLS only (n = 24) (0.135±0.137 mm2 and 3.338±2.929 mm2, respectively; p = 0.007). The BLS treated mice had significantly reduced survival compared to FGS- and FGS-UVC-treated mice for both relapse-free survival (RFS) (p

BACKGROUND:Refluxogenic effects of smoking and alcohol abuse may be related to the risk of esophageal squamous cell carcinoma (ESCC). The present study attempts to clarify the effects of continuous taurocholic acid (TCA) exposure, which is neither mutagenic nor genotoxic, on ESCC progression. METHODS:A squamous carcinoma cell line (ESCC-DR) was established from a tumor induced in a rat model of gastroduodenal reflux. ESCC-DR cells were incubated with 2 mM TCA for ≥2 months. The effects of continuous TCA exposure were evaluated in vitro on cell morphology, growth, and invasion and in vivo on xenograft tumor growth in nude mice. Moreover, the mean level of secreted transforming growth factor (TGF)-β1 and vascular endothelial growth factor (VEGF) proteins in cell culture supernatants and mRNA synthesis of TGF-β1 and VEGF-A of ESCC cells were measured. The angiogenic potential was further examined by a migration assay using human umbilical vein endothelial cells (HUVECs). RESULTS:Continuous TCA exposure induced marked formation of filopodia in vitro. Expression levels of angiogenic factors were significantly higher in the cells treated with TCA than in control cells. Tumor xenografts derived from cells pre-exposed to TCA were larger and more vascularized than those derived from control cells. In addition, TCA exposure increased HUVEC migration. CONCLUSION:Continuous TCA exposure enhanced ESCC progression due to reduced cell loss in vivo. Cell loss was inhibited by TCA-induced vascular endothelial cell migration, which was mediated by TGF-β1 and VEGF-A released from ESCC cells.

We aimed to clarify the association between a novel serum fibrosis marker, Wisteria floribunda agglutinin-positive Mac-2-binding protein (WFA+-M2BP), and hepatocellular carcinoma (HCC) development in 355 patients with chronic hepatitis C who achieved sustained virologic response (SVR) through interferon-based antiviral therapy. Pretreatment serum WFA+-M2BP levels were quantified and the hazard ratios (HRs) for HCC development were retrospectively analyzed by Cox proportional hazard analysis. During the median follow-up time of 2.9 years, 12 patients developed HCC. Multivariate analysis demonstrated that high serum WFA+-M2BP (≥2.80 cut off index (COI), HR = 15.20, p = 0.013) and high fibrosis-4 (FIB-4) index (≥3.7, HR = 5.62, p = 0.034) were independent risk factors for HCC development. The three- and five-year cumulative incidence of HCC in patients with low WFA+-M2BP were 0.4% and 0.4%, respectively, whereas those of patients with high WFA+-M2BP were 7.7% and 17.6%, respectively (p < 0.001). In addition, combination of serum WFA+-M2BP and FIB-4 indices successfully stratified the risk of HCC: the five-year cumulative incidences of HCC were 26.9%, 6.8%, and 0.0% in patients with both, either, and none of these risk factors, respectively (p < 0.001). In conclusion, pretreatment serum WFA+-M2BP level is a useful predictor for HCC development after achieving SVR.

Behavioral, anatomical, and gene expression studies have shown functional dissociations between the dorsal and ventral hippocampus with regard to their involvement in spatial cognition, emotion, and stress. In this study we examined the difference of the multisynaptic inputs to the dorsal and ventral dentate gyrus (DG) in the rat by using retrograde trans-synaptic tracing of recombinant rabies virus vectors. Three days after the vectors were injected into the dorsal or ventral DG, monosynaptic neuronal labeling was present in the entorhinal cortex, medial septum, diagonal band, and supramammillary nucleus, each of which is known to project to the DG directly. As in previous tracing studies, topographical patterns related to the dorsal and ventral DG were seen in these regions. Five days after infection, more of the neurons in these regions were labeled and labeled neurons were also seen in cortical and subcortical regions, including the piriform and medial prefrontal cortices, the endopiriform nucleus, the claustrum, the cortical amygdala, the medial raphe nucleus, the medial habenular nucleus, the interpeduncular nucleus, and the lateral septum. As in the monosynaptically labeled regions, a topographical distribution of labeled neurons was evident in most of these disynaptically labeled regions. These data indicate that the cortical and subcortical inputs to the dorsal and ventral DG are conveyed through parallel disynaptic pathways. This second-order input difference in the dorsal and ventral DG is likely to contribute to the functional differentiation of the hippocampus along the dorsoventral axis.

The glycoprotein (G) of rabies virus (RV) is required for binding to neuronal receptors and for viral entry. G-deleted RV vector is a powerful tool for investigating the organization and function of the neural circuits. It gives the investigator the ability to genetically target initial infection to particular neurons and to control trans-synaptic propagation. In this study we have quantitatively evaluated the effect of G gene deletion on the cytotoxicity and transgene expression level of the RV vector. We compared the characteristics of the propagation-competent RV vector (rHEP5.0-CVSG-mRFP) and the G-deleted RV vector (rHEP5.0-ΔG-mRFP), both of which are based on the attenuated HEP-Flury strain and express monomeric red fluorescent protein (mRFP) as a transgene. rHEP5.0-ΔG-mRFP showed lower cytotoxicity than rHEP5.0-CVSG-mRFP, and within 16 days of infection we found no change in the basic electrophysiological properties of neurons infected with the rHEP5.0-ΔG-mRFP. The mRFP expression level of rHEP5.0-ΔG-mRFP was much higher than that of rHEP5.0-CVSG-mRFP, and 3 days after infection the retrogradely infected neurons were clearly visualized by the expressed fluorescent protein without any staining. This may be due to the low cytotoxicity and/or the presumed change in the polymerase gene (L) expression level of the G-deleted RV vector. Although the mechanisms remains to be clarified, the results of this study indicate that deletion of the G gene greatly improves the usability of the RV vector for studying the organization and function of the neural circuits by decreasing the cytotoxicity and increasing the transgene expression level.

BACKGROUND: Ezrin is a member of the ezrin, radixin, and moesin family that provides a functional link between the plasma membrane and the cortical actin cytoskeleton. A correlation between ezrin overexpression and aggressive cancer behavior has been recently reported in various tumor types. However, its roles in the mechanisms underlying progression of tongue squamous cell carcinoma (SCC) are unclear. METHOD: We used human tongue SCC and noncancerous tissue microarrays to immunohistochemically analyze the ezrin expression level and its relationship with proliferative activity. The human tongue SCC cell line HSC-3 was used to determine the effects of ezrin RNA interference (RNAi) on cancer cells during MTT; wound healing and invasion assays; immunofluorescence of the actin cytoskeleton; and western blotting of E-cadherin, N-cadherin, β-catenin, and the active and total RhoA/Rac1/cdc42. RESULTS: Ezrin was overexpressed in 46.4% of the tumors examined in human tongue SCC tissue microarrays. Ezrin expression was correlated with the Ki-67 index. Ezrin depletion by RNAi in the HSC-3 cells significantly reduced cell proliferation, migration, and invasiveness and disturbed actin reorganization during podia formation. Its effects on RhoA/Rac1/cdc42 expression were not significant, whereas it enhanced E-cadherin and β-catenin expression and decreased N-cadherin expression. CONCLUSIONS: Ezrin is often overexpressed in primary tongue SCCs and may have an important role in their growth, migration, and invasiveness possibly via its relationship with the E-cadherin/β-catenin complex and the cadherin switch. Thus, ezrin could be a therapeutic target in tongue SCC.